Separation of Carbohydrate and Glycopeptide Isomers using Ion Mobility-Mass Spectrometry
Event details
| Date | 10.12.2015 |
| Hour | 16:30 › 17:30 |
| Speaker |
Prof. Kevin Pagel Freie Universität Berlin, Institute for Chemistry and Biochemistry and Fritz Haber Institute of the Max Planck Society, Berlin, Germany |
| Location | |
| Category | Conferences - Seminars |
Complex carbohydrates are of great importance for a variety of biological functions. They can be linked to proteins as post-translational modifications or exist as free oligosaccharides, for example in milk. Currently, the vast majority of complex carbohydrates are characterized using mass spectrometry-based techniques (MS) that are directly descending form those established in proteomics. Measuring the molecular weight of a sugar, however, immediately poses a fundamental problem: entire classes of the constituting monosaccharide building blocks exhibit an identical atomic composition and, consequently, also an identical mass. Therefore, carbohydrate MS data can be highly ambiguous and often it is simply not possible to clearly assign a particular molecular structure.
A promising approach to overcome the above-mentioned limitation is to implement an additional gas phase separation step using ion mobility-mass spectrometry (IM MS). Here, ions travel through a gas-filled cell aided by an electric field and are separated according to their mass, charge, size and shape. While the drift time of an ion depends on the underlying instrument conditions, the collision cross section (CCS) is a molecular property that can be compared and calculated theoretically.
Here, we demonstrate the potential of IM-MS to be used as a tool for the separation and identification of isomeric carbohydrates and glycopeptides. First, six synthetic carbohydrate isomers that differ with respect to their composition, connectivity, or configuration were analyzed individually and as mixtures. Our data reveal that carbohydrate linkage- and stereoisomers, which are difficult to distinguish using established techniques such as liquid chromatography or NMR, can be separated and unambiguously identified on basis of their CCS. When mixed, even minor isomeric components with relative concentrations as low as 0.1% were still clearly detectable.
Second, we recently extended our investigations to glycopeptides. First preliminary data show that the position of the carbohydrate within the peptide sequence can be clearly identified using IM-MS when multiple glycosylation sites are present. More importantly, however, our data reveal that also glycopeptides, which merely differ in the regiochemistry of the attached carbohydrate, can be distinguished using gas-phase fragmentation and subsequent IM-MS analysis.
These results indicate that IM-MS is an exceptionally effective tool for the structural analysis of complex carbohydrates. The full benefit of this method will become apparent once CCS data of carbohydrates and oligosaccharide fragments will be deposited in databases.
A promising approach to overcome the above-mentioned limitation is to implement an additional gas phase separation step using ion mobility-mass spectrometry (IM MS). Here, ions travel through a gas-filled cell aided by an electric field and are separated according to their mass, charge, size and shape. While the drift time of an ion depends on the underlying instrument conditions, the collision cross section (CCS) is a molecular property that can be compared and calculated theoretically.
Here, we demonstrate the potential of IM-MS to be used as a tool for the separation and identification of isomeric carbohydrates and glycopeptides. First, six synthetic carbohydrate isomers that differ with respect to their composition, connectivity, or configuration were analyzed individually and as mixtures. Our data reveal that carbohydrate linkage- and stereoisomers, which are difficult to distinguish using established techniques such as liquid chromatography or NMR, can be separated and unambiguously identified on basis of their CCS. When mixed, even minor isomeric components with relative concentrations as low as 0.1% were still clearly detectable.
Second, we recently extended our investigations to glycopeptides. First preliminary data show that the position of the carbohydrate within the peptide sequence can be clearly identified using IM-MS when multiple glycosylation sites are present. More importantly, however, our data reveal that also glycopeptides, which merely differ in the regiochemistry of the attached carbohydrate, can be distinguished using gas-phase fragmentation and subsequent IM-MS analysis.
These results indicate that IM-MS is an exceptionally effective tool for the structural analysis of complex carbohydrates. The full benefit of this method will become apparent once CCS data of carbohydrates and oligosaccharide fragments will be deposited in databases.
Practical information
- General public
- Free
Organizer
- ISIC Physical Chemistry Seminars
Contact
- Dr Frank van Mourik